For the last 20 years matrix metalloproteinases (MMPs) have been identified as important contributors to tumor progression on the basis of their abundant presence in neoplastic tissues, their ability to degrade all components of the extracellular matrix (ECM), and preclinical studies demonstrating the potent anti-tumor activity of MMP inhibitors (MPIs). In our laboratory and over the last 15 years of funding from this grant we have moved from an initial focus on MMP inhibition as a mechanism to prevent tumor invasion and metastasis to a focus on earlier stages of tumorigenesis. Our recent data in the MMTV-Wnt-1 de novo mouse mammary tumor model suggest that MMPs play a critical role in epithelial-mesenchymal transformation (EMT) and in tumor growth and angiogenesis. We hypothesize that there is a specific temporal expression of MMPs during Wnt-1 induced mammary tumorigenesis, with an initial dependence on neoplastic-derived MMP-3 for EMT and later dependence on MMP-9 for angiogenesis and tumor growth and that by targeting these two proteases, MPIs can inhibit tumorigenesis. This hypothesis is based on the observation that upon transformation with Wnt-1 mammary epithelial cells overexpress MMP-3, whereas Wnt-1 induced mammary tumors express MMP-9. In support of the hypothesis, we also observed that overexpression of a human tissue inhibitor of metalloproteinase-2 (TIMP- 2) transgene in the mammary gland of MMTV-Wnt-1 mice prevents or significantly delays the formation of tumors and results in slower growing tumors that are rich in collagen and contain less vessels. We propose 3 specific aims. The first aim will examine the function of MMP-3 in Wnt-1 mediated EMT, exploring whether MMP-3 is a downstream event in the Wnt-1 signaling pathway that is necessary or sufficient for transformation and determining the role of MMP-3 and the ECM in migration and invasion of Wnt-1 transformed mammary epithelial cells. The second aim will focus on examining the mechanisms of tumor inhibition by TIMP-2 and MPIs in MMTV-Wnt-1 mice, with particular emphasis on identifying the role of MMP- 3, MMP-9 and fibrillar collagen in tumor growth and angiogenesis, and on documenting proteolytic activity by imaging techniques. The third aim will determine whether MMPs are involved in tumor development in other de novo mammary tumor models that more closely mimic human breast cancer and in testing the role of MPIs in these models as chemoprevention. These studies will bring our understanding of MMP and MMP inhibition in cancer to a next level, and will generate information that could be critical in the design of better therapeutic or chemoprevention trials with MPIs. [unreadable] [unreadable]